Serotonin-6 (5-HT6) receptors are more heavily expressed in striatal medium spiny neurons than anywhere else in human or rat brain. Therefore, they are positioned to mediate a significant proportion of serotonin's effect on motivated behavior, such as is involved in learning and maintenance of self-administration of natural and drug rewards. We previously established that 5-HT6 receptors interfere with the acquisition of reward motivated learning in both dorsal and ventral striatum. Our core hypothesis is that, since 5-HT6 receptors are expressed in both the direct and indirect pathway medium spiny neurons, these receptors activate both pathways to a similar extent. This opposes the action of dopamine, which activates direct pathway neurons via D1 receptors and inhibits indirect pathway neurons via D2 receptors. Thus, dopamine turn on the "on" switch and off the "off" switch, and differential activity in these pathways promotes motivated behavior and procedural learning whereas 5-HT6 receptor activation reduces this differential activity by activating both pathways simultaneously. In order to test this hypothesis we have developed viral vectors that allow us to express transgenes selectively in the direct or indirect pathway medium spiny neurons. These vectors are based on the dynorphin or enkephalin promoter, which differentially target the direct and indirect pathway medium spiny neurons, respectively. We have collected strong evidence that the pDYN and pENK vectors target these pathways selectively. This provides us with key tools to increase or decrease 5-HT6 receptor expression in these pathways differentially and to test the cellular mechanism underlying striatal 5-HT6 receptor actions. The overall plan for this proposal is to use these vectors to disentangle the role of 5-HT6 receptors in direct and indirect pathway neurons of both dorsal and ventral striatum. We will use either overexpression or RNAi knockdown to modulate 5-HT6 receptor expression in either pathway to examine 5-HT6 receptors in discrete subregions of striatum on instrumental learning for a natural reward as well as a drug reward (cocaine). We will also examine the impact of these receptors in each pathway on motivation for cocaine once self-administration is established, and their effect on compulsive drug taking in the face of negative consequences to cocaine taking. The hypothesis, strategy, and tools involved in this project are all highly innovative. The long-term goal of this work is to understand the impact of serotonin on these distinct pathways in drug abuse so that better treatments for addiction can be developed. PUBLIC HEALTH RELEVANCE: Addiction is a behavioral and brain disease that involves adaptations that interfere with normal flexibility in behavior such that drugs are taken repeatedly and habitually despite negative consequences. We will use state of the art gene manipulations in targeted subgroups of neurons in the rat striatum to understand how 5- HT6 serotonin receptors participate in the acquisition and maintenance of reward motivated learning and cocaine self administration. The goal is to understand how serotonin participates in the adaptations associated with addiction so that we can design treatments to prevent or reverse these changes.